The invention relates to a method for the electron spin resonance (ESR)-spectroscopic determination of changes in the transport properties of albumin in an albumin-containing sample, which method is applicable in the medical, biological, biotechnological and veterinary practice and useful in diagnosing and/or monitoring physiological or pathological changes in a human or animal body, or in the quality control of albumin-containing preparations, particularly blood products. The invention is also directed to an ESR spectrometer for performing the method according to the invention.
The utilization of hematologic parameters or of parameters relating to antigens, hormones, enzymes, and other biologically active substances in the diagnosis of diseases is well-known and widely used in clinical-chemical routine diagnostics.
Similarly, NMR and ESR spectroscopy can be used in clinical diagnostics to detect structural and functional properties of protein and lipid components in blood serum. While the use of ESR spectroscopy employing spin labels to investigate the movement behavior in macromolecules represents the well-known state of the art, the extensive introduction of this testing method in clinical routine diagnostics has failed heretofore due to high cost of equipment and complicated or non-reliable analysis of the signals.
SU 1319705 A1 describes a method of diagnosing malignant diseases by measuring the ability of blood plasma or blood serum to bind an incorporated spin probe, using ESR. Therein, the binding coefficient α representing the ratio of the peak A amplitude to the peak B amplitude is determined. If α>1, a cancerous disease is present. If α<1, the subject is healthy.
The described method involves the disadvantage of operating with insufficient sensitivity, thus being incapable of providing a reliable diagnosis. Investigations on this method have shown the false-positive and false-negative diagnostic data at an early stage of disease to be ≧30%.
EP 0,973,043 A1 describes an ESR-spectroscopic method of diagnosing malignant neoplasms in blood serum, wherein physicochemical parameters of the mobility of a spin probe represented by a spin-labelled fatty acid are determined in the three binding sites of albumin.
Albumin is the main component in the transport system of blood, enabling the transport of fatty acids, tryptophane, bilirubin, calcium, steroid hormones, and other functionally significant active substances to the target cell, and being involved in binding and distribution of a variety of toxins (including those of endogenous origin). Albumin is a polypeptide (molecular weight 68,000) having a high binding capacity for exogenous (drugs) and endogenous substances. It is comprised of 585 amino acids arranged in a loop structure via disulfide bridges. The literature differentiates between distinct binding sites on serum albumin. For fatty acids, there are the so-called binding sites 1, 2 and 3 on the albumin molecule.
In the diagnostic method according to EP 0,973,043 A1, a cancerous disease is detected using the deviation of the parameters of spin probe binding from those values exhibited by healthy subjects.
Thus, the ESR method according to EP 0,973,043 A1 detects the concrete native conformational state of albumin in blood, which differs from that of a healthy subject in the presence of a cancerous pathology. This method has been found to produce false-positive or false-negative results in the presence of additional effects on albumin from the blood, e.g. elevated blood lipid values, ingestion of medicaments, or effects caused by the solvent of the spin probe or by storing the blood sample on air.